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学科主题: 分析化学
题名: Immobilized capillary enzyme reactor based on layer-by-layer assembling acetylcholinesterase for inhibitor screening by CE
其他题名: 基于层层自组装固定化毛细管酶微反应器用于乙酰胆碱酯酶的抑制剂筛选
作者: Tang ZM(唐忠美) ; Wang TD(汪彤丹) ; Kang JW(康经武)
通讯作者: 康经武
刊名: Electrophoresis
发表日期: 2007
卷: 28, 期:17, 页:2981-2987
收录类别: SCI
部门归属: 中科院上海有机化学研究所; 中科院研究生院
英文摘要: A method for creating an immobilized capillary acetylcholinesterase (AChE) reactor based on a layer-by-layer (LBL) assembly for inhibitor screening is described. The unique capillary AChE reactor was easily prepared by the instrument in three steps: first, a 0.5 cm long plug of a solution of the cationic polyelectrolyte polydiallyldimethylammonium. (PDDA) was injected into the capillary to produce a positively charged coating on the surface of the capillary; subsequently, the enzyme solution with the same plug length was injected into the capillary and incubated for 10 min to immobilize the enzyme on the capillary wall via electrostatic interaction; third, PDDA solution with the same plug length was injected again into the capillary to cover the immobilized enzyme by forming PDDA-AChE-PDDA sandwich-like structure. The enzyme reactor can be easily renewed after removing the immobilized enzyme by flushing the column with 1 M NaCl solution. Activity of the immobilized enzyme can be assayed simply by carrying out an electrophoretic separation, i.e., the substrate solution was injected and incubated for a short time, followed by applying a voltage to separate the product from the unreacted substrate. The measured peak area of the product then represented the enzyme activity. For enzyme inhibitor screening, the mixture solution of the substrate and the inhibitor was injected and assayed the reduction of the enzyme activity. The immobilized enzyme could withstand 100 consecutive assays by only losing 10% activity. The reproducibility in terms of time-to-time, day-to-day, and batch-to-batch was measured with RSD% less than 4.7%. Furthermore, the screening system was validated by a known inhibitor. Finally, screening a small compound library containing four known AChE inhibitors and 42 natural extracts was demonstrated, and species with inhibition activity can be straightforwardly identified with the system.
语种: 英语
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WOS记录号: WOS:000249743500005
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内容类型: 期刊论文
URI标识: http://ir.sioc.ac.cn/handle/331003/27673
Appears in Collections:上海有机化学研究所_期刊论文

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Recommended Citation:
Tang ZM,Wang TD,Kang JW. Immobilized capillary enzyme reactor based on layer-by-layer assembling acetylcholinesterase for inhibitor screening by CE[J]. Electrophoresis,2007,28(17):2981-2987.
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